Immobilized biocatalysts have found applications in a variety of industries where specific chemical conversions are required. Specific large scale examples in the food and pharmaceutical industries include immobilized glucose isomerase for the conversion of glucose to fructose and immobilized penicillin acylase for the preparation of derivatives of penicillin. Immobilized enzymes can be used for large scale bioremediation such as the destruction of undesirable chemical compounds. Another application involves small scale immobilized enzymes used in diagnostic reactions wherein the biocatalyst facilitates a chemical reaction which produces a detectable chemical moiety or other chemical change. More applications for immobilized enzymes will likely be developed in the future.
Immobilized biocatalysts (enzymes) provide advantages over bio catalysts in solution such as those provided in a liquid form or in a dry form to be dissolved in a liquid. Immobilized enzymes can be easily separated from the solution and thus permit the reuse of these enzymes hence reducing the cost of the enzyme in the production of a product. In the two cases cited above, immobilized enzymes are used in a packed column with the reactant solution pumped through the column and the desired chemical conversion accomplished when the solution leaves the column. In this process, the enzymes are used multiple times at high concentrations to achieve efficient usage of the enzymes.
In addition to ease of separation, immobilized enzymes can provide other potential advantages including greater thermal stability, pH stability, and the like during storage as well as during usage. Immobilized biocatalysts also can provide greater activity in various solvents compared to the enzymes in solution. There are often advantages to the physical form because immobilized biocatalysts can be provided in a granular form which reduces dust and improves handling of the product. As with a dried enzyme, the activity of the immobilized enzyme per mass of material can be varied as desired for a particular application.
An immobilized enzyme may be composed of the enzyme material combined with a solid matrix to which the enzyme is attached or held by chemical or physical means. The enzyme may be attached to the surface of the matrix, or if the matrix is porous enough to permit diffusion of the substrate, the enzyme may be distributed throughout the matrix. There are many methods proposed for the immobilization of biocatalysts including the entrapment of the enzyme in gel, the covalent, hydrophobic, electrostatic, and other methods for attachment of the enzyme to an inorganic or organic solid. The cross linking of the enzyme with whole cells, enzyme crystals, etc may be accomplished using reagents such as glutaraldehyde (GA) and polyethylenimine (PEI).
The conversion of a biocatalyst to an immobilized form may require more processing and hence may increase the cost of production for that enzyme form. Any increased cost must be compensated by increased productivity, shelf life, use in solvents, or some other advantage. Therefore, the cost of immobilization is an important consideration in the production of an immobilized enzyme. A lower cost for immobilization hence provides for a more economical usage of the immobilized enzyme.
Of the methods proposed for the immobilization of the enzymes, the use of cross linking whole cells is one of the most cost-effective. This method utilizes the whole cells are those of the microorganism used to make the enzyme and hence are available for no cost or even a negative cost if there is a cost associated with the disposal of the cell mass.
Various immobilization processes have been suggested for enzymes including attachment to silica supports. This process involves the fermentation of microorganism for the production of the enzyme, the separation (or purification) of the enzyme from the biomass, the chemical attachment or encapsulation of the enzyme to the matrix, and the production of the final product form. These immobilizations have the disadvantage of incurring the cost of the matrix to which the enzyme is attached. Further the separation of the enzyme from the fermentation biomass involves some loss of enzyme and enzyme activity, as well as associated purification processing costs. These disadvantages are overcome by the immobilization of the enzyme directly with the biomass used to produce the enzyme.
Some enzyme substrates have limited solubility in water and have better solubility in organic solvents or mixtures of organic solvents and water. For example, due to these properties, organic solvent solutions are often used to wash away organophosphorus compounds or organic solvents are used in the processing of triglycerides. Soluble enzymes are often deactivated or rendered inert in organic solvents or mixtures; however, immobilized enzymes can maintain some activity in solvents compared to soluble enzymes. Hence immobilized enzymes have the capacity to react with the higher concentrations of their substrates in such solvents.
Application of immobilized enzymes may require the disposal of the enzyme after usage. For example, the immobilized enzyme used in a column reactor would require disposal after use. Also, some immobilized enzymes can be considered for use directly in the environment. For example, immobilized OPH enzyme could be dispersed on an agricultural field for decontaminating a pesticide or other organophosphorus compound.